1. Field of the Invention
The subject invention is directed to a device and system that expedites and automates thawing of viably frozen cells, and more particularly, to an adaptor for supporting or otherwise suspending a cryovial containing a cryopreserved sample of viable cells over a centrifuge tube containing a cell culture medium.
2. Description of Related Art
The preservation of cells is an extremely important aspect of cell culture and fundamental to biological research. The only effective means of viably preserving eukaryotic cells is by freezing, also known as cryopreservation, that can be accomplished with either liquid nitrogen or by employing cryogenic freezers. The freezing process involves slowly reducing the temperature of prepared cells to −30° C. to −60° C. followed by a transfer to temperatures less than −130° C. Once at ultralow temperatures, the cells are biologically inert and can be preserved for years.
Cryopreserving eukaryotic cells differs from preserving bacteria and fungi in that higher viability is required. Where a 1% survival rate of a microbial culture can be practical, such low viability is unacceptable with cultured cells. High survival rates may be very important for cell lines due to the expense and difficulty in preparation, slow relative rate of growth, and tendency to change with repeated passage in culture. In addition, in many cases expansion of cryopreserved cells is not possible—for example, when working with cells isolated from blood or tissue specimens. Consequently, methods used for cell culture cryopreservation must ensure high viability (e.g., >80%).
Another important criterion is the overall yield (i.e., cells recovered after thawing compared to cells frozen). In particular, cells from tissue specimens are highly valuable and comprise a limiting resource. Maximizing yield of viably recovered cells is paramount.
Finally, the variability in these values must minimized. Differential viability and yield are known to impact assays performed following cell thawing. In particular, assays performed in GLP clinical laboratory settings place a high premium on minimizing the variability; in some settings, small decrements in viability or yield are tolerated to achieve lesser variation.
Factors that can affect the viability of cryopreserved cells include growth conditions prior to harvesting, the physiological state of the cells, the cell density, choice of cyroprotectant, and handling techniques. For cells isolated from tissues, there is no choice for most of these, and handling techniques become the principal source of variability. Cryoprotectants such as DMSO are valuable to prevent cell lysis during the freezing process. The diffusion of cryoprotective agents into a cell will result in a partial replacement of intracellular water and help to prevent dehydration (from ice formation) during freezing. Glycerol is also known to stabilize proteins in their native states and to assist in the maintenance of critical macromolecular interactions at subzero temperatures.
Nearly every cell biology laboratory will thaw cryopreserved specimens at some point in time, with large networks of vaccine or disease pathogenesis laboratories thawing thousands of cryopreserved cell specimens every year. In recent years, as technology has become available to measure more parameters from a single specimen, the size of immunological studies has grown greatly. A well-powered study can require tens of thousands of samples, and although high-throughput devices have been developed for automated acquisition of data, thawing cryopreserved specimens remains a labor-intensive, low-throughput endeavor.
The common method involves partially submerging cell vials in a 37° C. water bath, and then “swirling” the vials so that heat is evenly distributed, until only a “pea-sized” ice crystal remains. Because this is an intricate process, no more than four vials can be thawed at a time, severely limiting throughput. The subjective nature of the process also introduces problems, as the duration of the water bath step can vary dramatically from thaw to thaw, researcher to researcher, or by number of vials. This has important consequences for downstream assays, since incomplete thawing results in poor cell recovery, while excessive thawing time decreases cell viability. Additionally, the exposure of vials to an open 37° C. water bath increases the possibility of bacterial/fungal contamination.
There is a need for a system and method for expediting and automating thawing of cryopreserved specimens that overcomes the deficiencies and limitations of prior art systems and methods.